The primary goal of this shared instrumentation grant application is to develop multiphoton-based microscopic deep tissue intravital/ ex vivo/ in vivo/ i vitro imaging capability for basic, translational, pre-clinical and clinical research purposes at te Penn State Hershey College of Medicine. As evidenced by numerous peer-reviewed publications in leading scientific journals, the biomedical imaging of complex cellular level processes involved in various biological processes is a critical component of multiple research areas at the Penn State Hershey College of Medicine. Our investigators focus on understanding and elucidating complex intracellular processes in many areas, including immunology, infectious diseases, cancer, neurodegenerative diseases, behavioral and neuroscience, kidney disease, and lung disease. For visualizing relevant and complex extra/ intra-cellular dynamic processes deep within the tissues in vivo at high spatial (submicron) and temporal (submillisecond) resolutions, a high resolution deep tissue imaging tool with ultra-high speed scanning such as the Nikon Nikon A1 MP+ Multiphoton Microscope is required. Multimodal multiphoton microscopy has become a powerful imaging method for intravital/ ex vivo/ in vivo/ in vitro evaluation of extra/ intracellular structures deep within tissues in their native environments. Thi technology, which uses femto-second Infrared (IR) laser pulses as the excitation source, is efficient in producing multiphoton excitation fluorescence (MPEF) from endogenous/exogenous fluorescent macromolecules, and inducing highly specific second harmonic generation (SHG) signals from non-centrosymmetric biomolecules such as fibrillar collagens, myosins and microtubules. For instance, these multimodal high resolution imaging approaches can be an optimal tool to visualize and quantify the cellular dynamics of the immune response in lymphoid organs and in peripheral tissues; direct observation neuronal activity, morphological changes, cellular interactions in the intact and awake/behaving brain in most layers of the cortex including white matter and hippocampus; direct visualization of dynamic intra renal processes in real-time, extent of fibrosis or collagen fibrillation, cellular interactions in small animal models of cancer diseases, blood flow limitations as a result of cerebral malarial infections, live cell migrations, drug delivery and therapeutic developments among others. These imaging modalities are minimally invasive since structures deep within tissues can be visualized without the need for tissue fixation or sectioning. In summary, this multimodal system will enable us to add a crucial service in core facilities, which will be used by numerous NIH-funded investigators in addition to other well-funded research projects.